Live-cell imaging studies of central nervous system synapse formation and alteration.

中枢神经系统突触形成和改变的活细胞成像研究。

• 批准号: 17590152
• 负责人: INOUE Akihiro
• 金额: $ 1.86万
• 依托单位: Tokyo Medical and Dental University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2005
• 资助国家: 日本
• 起止时间: 2005 至 2006
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-17590152/
• 关键词: synapse scaffolding protein; virus vector; RNA interference; green fluorescent protein; post synaptic density; actin; hippocampal culture; fluorescent recovery after photobleaching; シナプス形成; トランスジェニックマウス; 脳組織器官培養標本; 二光子レーザー顕微鏡

Organization and dynamic remodeling of postsynaptic density (PSD) are thought to be important for postsynaptic signal transduction. To study the molecular mechanisms, we performed live-cell imaging analyses of dynamic characteristics of major PSD proteins both at steady state and when neurons are activated by exogenous stimulations.Cortactin is an F-actin-associated protein which interacts with the postsynaptic scaffolding protein Shank and is localized within the dendritic spine. Green fluorescent protein (GFP)-based time-lapse imaging revealed cortactin redistribution from dendritic cytoplasm to postsynaptic sites by application of brain-derived neurotrophic factor (BDNF). In contrast, activation of N-methyl-d-aspartate (NMDA) receptors induced loss of cortactin from postsynaptic sites. These results indicate the importance of counterbalance between BDNF and NMDA receptor-mediated signaling in the reorganization of the postsynaptic actin cytoskeleton during neuronal development.We also reported that four major scaffolding molecules, PSD-95, GKAP, Shank, and PSD-Zip45, show distinct instability in total molecular content per synapse. Fluorescence recovery after photobleaching (FRAP) also confirmed their distinct turnover rates. Among the PSD molecules examined, PSD-95 was most stable. Acute pharmacological disruption of F-actin eliminated the dynamic fraction of GKAP, Shank, and PSD-Zip45. GKAP content in synapses increased after pharmacological enhancement of neuronal activity, on the other hand Shank and PSD-Zip45 content are reduced. Inhibition of F-actin dynamics prevented activity-dependent redistribution of all three scaffolds. These results indicate that F-actin plays a key role for regulating the dynamic reorganization molecular composition in PSD.

突触后密度（PSD）的组织和动态重构被认为是突触后信号转导的重要因素。为了研究分子机制，我们对稳态和外源刺激激活神经元时主要PSD蛋白的动态特性进行了活细胞成像分析。皮质蛋白是一种与f -肌动蛋白相关的蛋白，它与突触后支架蛋白Shank相互作用，并定位于树突脊柱内。利用脑源性神经营养因子（BDNF），基于绿色荧光蛋白（GFP）的延时成像揭示了树突状细胞质皮层向突触后部位的再分布。相反，n -甲基-d-天冬氨酸（NMDA）受体的激活诱导突触后位点的接触丧失。这些结果表明，在神经元发育过程中，BDNF和NMDA受体介导的信号传导平衡在突触后肌动蛋白细胞骨架重组中的重要性。我们还报道了四个主要的支架分子，PSD-95， GKAP， Shank和PSD-Zip45，在每个突触的总分子含量上表现出明显的不稳定性。光漂白后荧光恢复（FRAP）也证实了它们不同的周转率。在所检测的PSD分子中，PSD-95最稳定。f -肌动蛋白的急性药理破坏消除了GKAP， Shank和PSD-Zip45的动态部分。药物增强神经元活性后突触内GKAP含量增加，而Shank和PSD-Zip45含量降低。抑制f -肌动蛋白动力学可阻止所有三种支架的活性依赖性再分布。这些结果表明，f -肌动蛋白在PSD的动态重组分子组成调控中起着关键作用。

项目成果

Direct visualization of cell movement in the embryonic olfactory bulb using green fluorescent protein transgenic mice : evidence for rapid tangential migration of neural cell precursors.

使用绿色荧光蛋白转基因小鼠直接观察胚胎嗅球中的细胞运动：神经细胞前体快速切向迁移的证据。

• 发表时间: 2005
• 期刊: Neuroscience Research 51
• 作者: Yamamoto;K.;Yamaguchi;M.;S.Okabe
• 通讯作者: S.Okabe

Telencephalin slows spine maturation

• DOI: 10.1523/jneurosci.2651-05.2006
• 发表时间: 2006-02-08
• 期刊: JOURNAL OF NEUROSCIENCE
• 影响因子: 5.3
• 作者: Matsuno, H;Okabe, S;Yoshihara, Y
• 通讯作者: Yoshihara, Y

Simultaneous Observation of Stably associated presynaptic varicosities and postsynaptic spine : morphological alteration of CA3-CA1.

同时观察稳定相关的突触前静脉曲张和突触后棘：CA3-CA1 的形态改变。

• 发表时间: 2005
• 期刊: Molecular Cellular Neuroscience 28
• 作者: Umeda;T. et al.
• 通讯作者: T. et al.

Bi-directional regulation of postsynaptic cortactin distribution by BDNF and NMDA receptor activity

• DOI: 10.1111/j.1460-9568.2005.04510.x
• 发表时间: 2005-12-01
• 期刊: EUROPEAN JOURNAL OF NEUROSCIENCE
• 影响因子: 3.4
• 作者: Iki, J;Inoue, A;Okabe, S
• 通讯作者: Okabe, S

Simultaneous observation of stably associated presynaptic varicosities and postsynaptic spines : morphological alterations of CA3-CA1 synapses in hippocampal slice cultures.

同时观察稳定相关的突触前静脉曲张和突触后棘：海马切片培养物中 CA3-CA1 突触的形态变化。

• 发表时间: 2005
• 期刊: Molecular and Cellular Neuroscience 28
• 作者: Umeda;T.;Ebihara;T.;S.Okabe
• 通讯作者: S.Okabe